1. Field of the Invention
The present invention relates to an assay method for measuring immune reactions and to its apparatus, which is used to detect a very small amount of organic material.
2. Discussion of Background
As immunological means used for detecting a very small amount of organic material, a great number of studies and propositions on assay methods for detecting immune reactions have been recently made, including radioimmunoassay, fluorescent antibody method, enzyme antibody method, enzymeimmunoassay, etc.
In these representative test methods, either an antigen or an antibody (or anti-immunoglobulin), which undertakes a specific reaction, is conjugated with an appropriate marker, the complex of the antigen-antibody-marker or antibody-antigen-marker is formed, and the amount of the complex is quantitatively detected by measuring the marker as a criterion. The complex is formed by binding to the surface of a cell on which either an antigen or an antibody is fixed, or to the surface of a bead (particle) which is charged into a cell and on which either an antigen or antibody is fixed.
In other methods excepta radioimmunoassay using a radioactive substance as a marker, a fluorescent substance or a luminescent substance used as a marker is directly detected optically, or the catalytic activity of an enzyme used as a marker is revealed by the addition of an appropriate substrate and absorbance, fluorescence or luminescence is determined optically.
In various means for optical determination, the principle is that a microtiter plate serving as a cell or a sample reaction chamber, or a reactor (referred to as a cell hereinafter) composed of a separate test cup is placed opposite to the optical system, which receives reflected light from the cell, for use in determination of the intensity of light.
Since the amount of samples to be tested in immune reactions is extremelysmall, usually less than 10.sup.-13 mol/1, factors causing technical errors in the test should be excluded as much as possible even if precise equipment is employed in the aforesaid optical system.
The present inventors made various investigations from these points of view, and have learned that in the test method using a bead, which has been known as a method for detecting the immune reaction, beads existing in the cell affected technical errors in the test; that is, since the measuring area in the optical system which receives the reflected light from the cell is, in general, limit as to the measuring are within the cell, whether or not the bead exists within the measuring are of the optical system is considered to have an influence on measurement.